Project Summary Previously, we identified the small, Ras-like GTPase Rem2 as a critical regulator of neuronal morphology, synapse formation and neuronal plasticity in the vertebrate nervous system. Rem2 is a member of the RGK family of non-canonical Ras-like GTPases and is primarily expressed in the brain. Our published studies demonstrated that Rem2 functions in a CaMK signaling pathway that restricts dendritic branching. In fact, Rem2 is itself a substrate of CaMKII, and phosphorylation of Rem2 by CaMKII is required for Rem2 to regulate dendritic branching. CaMKII is an abundant protein kinase that serves many functions in diverse tissues including regulation of activity-dependent dendritic remodeling, Long Term Potentiation (LTP), which is the biological correlate of learning and memory, and regulation of hypertrophy and Ca2+ homeostasis in heart muscle. To better understand Rem2 signaling, we took an unbiased, proteomics approach to identify Rem2 interacting proteins and found that Rem2 interacts with all four CaMKII isoforms. Using an in vitro kinase assay with purified proteins, we demonstrated that Rem2, while a substrate of CaMKII, also potently inhibits its kinase activity. This suggests that Rem2 is a direct, endogenous inhibitor of CaMKII activity, a previously un-described function for this protein. While many pathways that activate CaMKII have been identified, there is only one other molecule described thus far in mammalian cells that inhibits CaMKII signaling (CaMKIIN). Interestingly, other RGK family members influence CaMKII activity in different cell types, although the exact mechanism of this regulation has not been reported. We propose experiments that will provide a detailed, mechanistic understanding of CaMKII inhibition by Rem2 and further, determine if inhibition of CaMKII activity is a conserved function of the RGK family.